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- Title
Nanobody-enabled monitoring of kappa opioid receptor states.
- Authors
Che, Tao; English, Justin; Krumm, Brian E.; Kim, Kuglae; Pardon, Els; Olsen, Reid H. J.; Wang, Sheng; Zhang, Shicheng; Diberto, Jeffrey F.; Sciaky, Noah; Carroll, F. Ivy; Steyaert, Jan; Wacker, Daniel; Roth, Bryan L.
- Abstract
Recent studies show that GPCRs rapidly interconvert between multiple states although our ability to interrogate, monitor and visualize them is limited by a relative lack of suitable tools. We previously reported two nanobodies (Nb39 and Nb6) that stabilize distinct ligand- and efficacy-delimited conformations of the kappa opioid receptor. Here, we demonstrate via X-ray crystallography a nanobody-targeted allosteric binding site by which Nb6 stabilizes a ligand-dependent inactive state. As Nb39 stabilizes an active-like state, we show how these two state-dependent nanobodies can provide real-time reporting of ligand stabilized states in cells in situ. Significantly, we demonstrate that chimeric GPCRs can be created with engineered nanobody binding sites to report ligand-stabilized states. Our results provide both insights regarding potential mechanisms for allosterically modulating KOR with nanobodies and a tool for reporting the real-time, in situ dynamic range of GPCR activity. Recent studies revealed that G protein-coupled receptors rapidly interconvert between multiple states. Here, authors use the kappa opioid receptor (KOR) and show how two state-dependent nanobodies provide real-time reporting of ligand stabilized states with KOR and other GPCRs.
- Subjects
OPIOID receptors; BINDING sites; G protein coupled receptors; X-ray crystallography; ALLOSTERIC regulation
- Publication
Nature Communications, 2020, Vol 11, Issue 1, p1
- ISSN
2041-1723
- Publication type
Article
- DOI
10.1038/s41467-020-14889-7